Poly-substituted-phenyl-oligoribo nucleotides having enhanced stability and membrane permeability and methods of use
In accordance with the present invention, antisense oligonucleotides are provided with enhanced membrane permeability and stability. This is accomplished in accordance with the invention through conjugating oligoribonucleotides with a hydrophobic carrier agent at the 2'-O position of the oligonucleotides. The hydrophobic carrier agent comprises a compound of the following general structure: ##STR1## wherein R.sup.1, R.sup.2, R.sup.3, R.sup.4, and R.sup.5 are independently H, NO.sub.2, halide, linear or branched alkyl, linear or branched acyl, linear or branched alkylene, linear or branched O-alkyl, linear or branched amido, linear or branched S-alkyl, mono or disubstituted amine, linear or branched thioamido, phosphothionate, or phosphothioate.In a preferred embodiment, R.sup.1 and R.sup.3 are NO.sub.2. In such embodiment, it will be appreciated that when R.sup.2, R.sup.4, and R.sup.5 are H, the compound is DNP and when R.sup.4 is F, the compound is FDNP.
Claims
1. An antisense oligoribonucleotide conjugated at the 2'-O position with a substituted phenyl group to produce a derivatized compound of the following general structure: ##STR18## wherein R.sup.1, R.sup.2, R.sup.3, R.sup.4, and R.sup.5 are independently H, NO.sub.2, halide, linear or branched alkyl, linear or branched acyl, linear or branched alkylene, linear or branched O-alkyl, linear or branched amido, linear or branched S-alkyl, mono or disubstituted amine, linear or branched thioamido, phosphorothioate, or phosphothioate.
2. The oligoribonucleotide of claim 1, wherein R.sup.1 and R.sup.3 are NO.sub.2 as follows: ##STR19##
3. The oligoribonucleotide of claim 2, wherein R.sup.2, R.sup.4, and R.sup.5 are H.
4. The oligoribonucleotide of claim 1, wherein the oligoribonucleotide has a length of between 10 and 40 nucleotides.
5. The oligoribonucleotide of claim 1, wherein the oligoribonucleotide has a length of between 12 and 30 nucleotides.
6. The oligoribonucleotide of claim 1, wherein the oligoribonucleotide has a length of between 15 and 25 nucleotides.
7. The oligoribonucleotide of claim 2, wherein the oligoribonucleotide has a length of between 10 and 40 nucleotides.
8. The oligoribonucleotide of claim 2, wherein the oligoribonucleotide has a length of between 12 and 30 nucleotides.
9. The oligoribonucleotide of claim 2, wherein the oligoribonucleotide has a length of between 15 and 25 nucleotides.
10. The oligoribonucleotide of claim 3, wherein the oligoribonucleotide has a length of between 10 and 40 nucleotides.
11. The oligoribonucleotide of claim 3, wherein the oligoribonucleotide has a length of between 12 and 30 nucleotides.
12. The oligoribonucleotide of claim 3, wherein the oligoribonucleotide has a length of between 15 and 25 nucleotides.
13. The antisense oligoribonucleotide of claim 1, wherein at least one of the 5'-OH and 3'-OH ends of the oligoribonucleotide is conjugated with a substituted phenyl group at a 2'-O ribose position to produce a derivatized compound of the following general structure: ##STR20## wherein R.sup.1, R.sup.2, R.sup.3, R.sup.4, and R.sup.5 are independently H, NO.sub.2, halide, linear or branched alkyl, linear or branched acyl, linear or branched alkylene, linear or branched O-alkyl, linear or branched amido, linear or branched S-alkyl, mono or disubstituted amine, linear or branched thioamido, phosphorothioate, or phosphothioate.
14. The oligoribonucleotide of claim 13, wherein R.sup.1 and R.sup.3 are NO.sub.2.
15. The oligoribonucleotide of claim 14, wherein R.sup.2, R.sup.4, and R.sup.5 are H.
16. An antisense oligoribonucleotide derivatized at a plurality of 2'-O positions with a hydrophobic group selected from the group consisting of a 2,4-dinitrophenyl group and a 3-fluoro-4,6-dinitrophenyl group.
17. The oligoribonucleotide of claim 16, wherein the oligoribonucleotide has a length of between 10 and 40 nucleotides.
18. The oligoribonucleotide of claim 16, wherein the oligoribonucleotide has a length of between 12 and 30 nucleotides.
19. The oligoribonucleotide of claim 16, wherein the oligoribonucleotide has a length of between 15 and 25 nucleotides.
20. A method of enhancing membrane permeability and stability of an oligoribonucleotide, comprising:
- providing an oligoribonucleotide having a plurality of 2'-O positions;
- conjugating a substituted phenyl group at said plurality of 2'-O ribose positions to produce a derivatized compound of the following general structure: ##STR21## wherein R.sup.1, R.sup.2, R.sup.3, R.sup.4, and R.sup.5 are independently H, NO.sub.2, halide, linear or branched alkyl, linear or branched acyl, linear or branched alkylene, linear or branched O-alkyl, linear or branched amido, linear or branched S-alkyl, mono or disubstituted amine, linear or branched thioamido, phosphorothioate, or phosphorothioate.
21. The method claim 20, wherein R.sup.1 and R.sup.3 are NO.sub.2 as follows: ##STR22##
22. The method claim 21, wherein R.sup.2, R.sup.4, and R.sup.5 are H.
23. The method of claim 20, wherein the oligoribonucleotide has a length of between 10 and 40 nucleotides.
24. The method of claim 20, wherein the oligoribonucleotide has a length of between 12 and 30 nucleotides.
25. The method of claim 20, wherein the oligoribonucleotide has a length of between 15 and 25 nucleotides.
26. The method of claim 21, wherein the oligoribonucleotide has a length of between 10 and 40 nucleotides.
27. The method of claim 21, wherein the oligoribonucleotide has a length of between 12 and 30 nucleotides.
28. The method of claim 21, wherein the oligoribonucleotide has a length of between 15 and 25 nucleotides.
29. The method of claim 22, wherein the oligoribonucleotide has a length of between 10 and 40 nucleotides.
30. The method of claim 22, wherein the oligoribonucleotide has a length of between 12 and 30 nucleotides.
31. The method of claim 22, wherein the oligoribonucleotide has a length of between 15 and 25 nucleotides.
32. The method of claim 20, wherein at least one of the 5'-OH and 3'-OH ends of the oligoribonucleotide is conjugated with a substituted phenyl group at a 2'-O ribose position to produce a derivatized compound of the following general structure: ##STR23## wherein R.sup.1, R.sup.2, R.sup.3, R.sup.4, and R.sup.5 are independently H, NO.sub.2, halide, linear or branched alkyl, linear or branched acyl, linear or branched alkylene, linear or branched O-alkyl, linear or branched amido, linear or branched S-alkyl, mono or disubstituted amine, linear or branched thioamido, phosphorothioate, or phosphorothioate.
33. The method of claim 32, wherein R.sup.1 and R.sup.3 are NO.sub.2.
34. The method of claim 33, wherein R.sup.2, R.sup.4, and R.sup.5 are H.
35. In an antisense therapeutic method, comprising administering an antisense oligonucleotide comprising a plurality of 2'-O positions in a manner designed inhibit gene expression, the improvement comprising:
- derivatizing the antisense oligonucleotide at a plurality of the 2'-O positions with a substituted phenyl group to produce a derivatized compound of the following structure: ##STR24## wherein R.sup.1, R.sup.2, R.sup.3, R.sup.4, and R.sup.5 are independently H, NO.sub.2, halide, linear or branched alkyl, linear or branched acyl, linear or branched alkylene, linear or branched O-alkyl, linear or branched amido, linear or branched S-alkyl, mono or disubstituted amine, linear or branched thioamido, phosphothionate, or phosphorothioate.
36. In the improvement of claim 35, wherein R.sup.1 and R.sup.3 are NO.sub.2 as follows: ##STR25##
37. In the improvement of claim 36, wherein R.sup.2, R.sup.4, and R.sup.5 are H.
38. In the improvement of claim 35, wherein the oligoribonucleotide has a length of between 10 and 40 nucleotides.
39. In the improvement of claim 35, wherein the oligoribonucleotide has a length of between 12 and 30 nucleotides.
40. In the improvement of claim 35, wherein the oligoribonucleotide has a length of between 15 and 25 nucleotides.
41. In the improvement of claim 36, wherein the oligoribonucleotide has a length of between 10 and 40 nucleotides.
42. In the improvement of claim 36, wherein the oligoribonucleotide has a length of between 12 and 30 nucleotides.
43. In the improvement of claim 36, wherein the oligoribonucleotide has a length of between 15 and 25 nucleotides.
44. In the improvement of claim 37, wherein the oligoribonucleotide has a length of between 10 and 40 nucleotides.
45. In the improvement of claim 37, wherein the oligoribonucleotide has a length of between 12 and 30 nucleotides.
46. In the improvement of claim 37, wherein the oligoribonucleotide has a length of between 15 and 25 nucleotides.
47. In the improvement of claim 35, wherein at least one of the 5'-OH and 3'-OH ends of the oligoribonucleotide is conjugated to a substituted phenyl group at a 2'-O ribose position to produce a derivatized with a compound of the following general structure: ##STR26## wherein R.sup.1, R.sup.2, R.sup.3, R.sup.4, and R.sup.5 are independently H, NO.sub.2, halide, linear or branched alkyl, linear or branched acyl, linear or branched alkylene, linear or branched O-alkyl, linear or branched amido, linear or branched S-alkyl, mono or disubstituted amine, linear or branched thioamido, phosphothionate, or phosphorothioate.
48. In the improvement of claim 47, wherein R.sup.1 and R.sup.3 are NO.sub.2.
49. In the improvement of claim 48, wherein R.sup.2, R.sup.4, and R.sup.5 are H.
50. In a method for administering an oligoribonucleotide, wherein the oligoribonucleotide comprises a plurality of 2'-O positions in a manner designed to inhibit gene expression the improvement comprising:
- derivatizing the O at a plurality of the 2'-O positions with a moiety selected from the group consisting of a 2,4-dinitrophenyl moiety and a 3-fluoro-4,6-dinitrophenyl moiety.
51. In the improvement of claim 50, wherein the oligoribonucleotide has a length of between 10 and 40 nucleotides.
52. In the improvement of claim 50, wherein the oligoribonucleotide has a length of between 12 and 30 nucleotides.
53. In the improvement of claim 50, wherein the oligoribonucleotide has a length of between 15 and 25 nucleotides.
54. In the improvement of claim 50, wherein at least one of the 5'-OH and 3'-OH ends of the oligoribonucleotide is conjugated with a substituted phenyl group at a 2'-O ribose position to produce a derivatized compound of the following general structure: ##STR27## wherein R.sup.1, R.sup.2, R.sup.3, R.sup.4, and R.sup.5 are independently H, NO.sub.2, halide, linear or branched alkyl, linear or branched acyl, linear or branched alkylene, linear or branched O-alkyl, linear or branched amido, linear or branched S-alkyl, mono or disubstituted amine, linear or branched thioamido, phosphorothioate, or phosphorothioate.
55. In the improvement of claim 50, wherein R.sup.1 and R.sup.3 are NO.sub.2.
56. In the improvement of claim 50, wherein R.sup.2, R.sup.4, and R.sup.5 are H.
57. The oligonucleotide of any of claims 1-56, wherein the oligonucleotide is an antisense oligonucleotide.
58. A method of derivatizing an oligoribonucleotide, comprising:
- providing an oligoribonucleotide having a plurality of 2'-O positions;
- conjugating a substituted phenyl group at a plurality of 2'-O positions to produce a derivatized compound of the following general structure: ##STR28## wherein R.sup.1, R.sup.2, R.sup.3, R.sup.4, and R.sup.5 are independently H, NO.sub.2, halide, linear or branched alkyl, linear or branched acyl, linear or branched alkylene, linear or branched O-alkyl, linear or branched amido, linear or branched S-alkyl, mono or disubstituted amine, linear or branched thioamido, phosphothionate, or phosphorothioate.
59. The method of claim 58, wherein R.sup.1 and R.sup.3 are NO.sub.2.
60. The method of claim 58, wherein R.sup.2, R.sup.4, and R.sup.5 are H.
61. A method of derivatizing an oligoribonucleotide, comprising:
- providing an oligoribonucleotide having a plurality of 2'-O positions;
- reacting said oligoribonucleotide with a Sanger-type reagent at a plurality of 2'-O positions to produce a derivatized oligoribonucleotide of the following general structure: ##STR29##
62. The method of claim 61, wherein R.sup.1 and R.sup.3 are NO.sub.2.
63. The method of claim 61, wherein R.sup.2, R.sup.4, and R.sup.5 are H.
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Type: Grant
Filed: Feb 22, 1996
Date of Patent: Jan 12, 1999
Inventor: Jui H. Wang (Amherst, NY)
Primary Examiner: John L. LeGuyader
Law Firm: Fliesler, Dubb, Meyer & Lovejoy
Application Number: 8/604,871
International Classification: A61K 4800; C12Q 168; C07H 2101;
